A simple RP-HPLC method for the simultaneous determination of citrulline and arginine in Australian cucurbits

Citrulline is a non-proteinaceous amino acid found in high concentrations in melons and other cucurbits. Via its subsequent biotransformation into arginine by the kidneys, citrulline plays important roles in nitric oxide production, vasodilation and ammonia detoxification. Dietary citrulline supplementation provides a range of benefits, including improved endothelial and cardiovascular function, and enhanced exercise capacity and post-exercise recovery. Hence there is considerable interest in developing citrulline-rich food products from melon cultivars containing high citrulline contents. However, most current methods for citrulline and arginine quantification require time-consuming chemical derivatization steps and use higher-end detection methods such as fluorescence detection. We report a simple, rapid reversed-phase high performance liquid chromatography method using ultraviolet detection (RP-HPLC-UV), without requiring analyte derivatization. Following optimisation of the column type, mobile phase composition, flow rate and detection wavelength, the method showed good linearity (R2=0.9995), a low limit of detection (0.6 and 0.3 mg L-1 for citrulline and arginine, respectively) and high repeatability (0.2% relative standard deviation). The optimised method was used to quantify the citrulline and arginine content of 13 Central Queensland fruit and vegetable cultivars, primarily cucurbits. The highest citrulline content and citrulline:arginine ratio were found in mature colocynth (Citrullus colocynthis) fruit (39.2 mg/g DWB), while red watermelon (Citrullus lanatus) flesh also contained high levels of citrulline (22.9 mg/g) and arginine (8.1 mg/g). Lower concentrations of citrulline and arginine were found in other melon cultivars, pumpkins and squash samples. This HPLC method may find use for the rapid screening of citrulline and arginine contents.